Studies in this laboratory concerning the hormonal regulation of gluconeogenesis have demonstrated a hormonal regulation of rat hepatic pyruvate kinase involving a reversible interconversion between two kinetically distinct forms of the enzyme. Our results are consistent with the hypothesis that pyruvate kinase is regulated by a cyclic-AMP dependent phosphorylation mechanism. The proposed investigation will include an examination of the various factors participating in the regulation of pyruvate kinese in the isolated rat hepatocyte with emphasis on determining the role of this enzyme in the physiological response of the liver to changing nutritional demands of the body. The studies will include a demonstration that the regulation of pyruvate kinase in the isolated hepatocyte by glucagon involves phosphorylation of the enzyme. The role of cyclic-AMP in the regulation of the enzyme by L-epinephrine and insulin will also be investigated. The enzymes involved in the phosphorylation and dephosphorylation of pyruvate kinase will be examined in the isolated hepatocyte and the influence of the allosteric effectors of pyruvate kinase (fructose-1,6-bisphosphate, ATP, and L-alanine) on these reactions will be characterized.